Fig. 6: Mutagenesis studies.

a Active site cavity of SptF with 1 (shown as cyan sticks). Hydrophobic residues (shown as orange sticks), Ile63, Phe133, Ile231, and V264’, form a hydrophobic surface to interact with the hydrophobic part of the substrate, whereas hydrophilic residues (shown as salmon sticks), Asn65, Ser114, Thr148, and Asn150, interact with the hydrophilic part of the substrate. b, c LC-MS EICs of products from in vitro assays with mutants using 1 or 15 as the substrate. The + m/z value used for each compound is shown. d Structures of new products generated by mutants. Compounds 32 and 33 were isolated from large scale enzymatic reactions using the I231A and F133A mutants, respectively. Structures were determined by NMR. e Ratios of 4 and 5 from mutants compared with those from wild-type SptF. All reactions were performed in triplicates. Data are presented as mean values and the error bars indicate standard deviations (SD). f Active site residues interacting with 1 (shown as cyan sticks), including hydrogen bonding directly with Ser114 and the newly introduced Thr65, and indirectly with Thr148 and Leu199 via waters in N65T (shown in purple). Note that the coloring scheme for ligands is the same as in Fig. 4.